Template polymerization initial reaction rate

Spontaneous polymerization of 4-vinyl pyridine in the presence of polyacids was one of the earliest cases of template polymerization studied. Vinyl pyridine polymerizes without an additional initiator in the presence of both low molecular weight acids and polyacids such as poly(acrylic acid), poly(methacrylic acid), polyCvinyl phosphonic acid), or poly(styrene sulfonic acid). The polyacids, in comparison with low molecular weight acids, support much higher initial rates of polymerization and lead to different kinetic equations. The authors suggested that the reaction was initiated by zwitterions. The chain reaction mechanism includes anion addition to activated double bonds of quaternary salt molecules of 4-vinylpyridine, then propagation in the activated center, and termination of the growing center by protonization. The proposed structure of the product, obtained in the case of poly(acrylic acid), used as a template is ... 

The kinetics of template polymerization depends, in the first place, on the type of polyreaction involved in polymer formation. The polycondensation process description is based on the Flory s assumptions which lead to a simple (in most cases of the second order), classic equation. The kinetics of addition polymerization is based on a well known scheme, in which classical rate equations are applied to the elementary processes (initiation, propagation, and termination), according to the general concept of chain reactions. 

Application of classical type of kinetic equations to the template polymerization was demonstrated by Kabanov at al It was shown that 4-vinylpyridine, in the presence of poly(methacrylic acid), poly(acrylic acid), poly(l-glutamic acid), and polyphosphate, polymerizes according to the classical equation and the order of reaction with respect to the monomer is 2 as demonstrated in the Figure 8.1. In log-log coordinates, for the all sets of polymerizations, experimental points fit straight lines. In the same paper dependence of the initial rate on the molar ratio of acid to monomer was examined. This relationship is shown on the Figure 8.2. The rate of polymerization in the presence of the poly(acrylic acid) is much higher than that for the low molecular analogue (acetic acid). The polymerization rate riches its maximum for the molar ratio [acid]/[monomer] 2. The authors found kinetic equation for template polymerization of 4-vinylpyridine in the presence of different polyacids in the form ... 

Description of polymerization kinetics in heterogeneous systems is complicated, even more so given that the structure of complex formed is not very well defined. In template polymerization we can expect that local concentration of the monomer (and/or initiator) can be different when compared with polymerization in the blank system. Specific sorption of the monomer by macromolecular coil leads to the increase in the concentration of the monomer inside the coil, changing the rate of polymerization. It is a problem of definition as to whether we can call such a polymerization a template reaction, if monomer is randomly distributed in the coil on the molecular level but not ordered by the template. 

Determination of the reaction rate from calorimetric measurements, using DSC technique, is very useful and was applied with success for many template polymerization systems and for blank polymerizations.Two types of calorimetric measurements were described isothermal and scanning experiments. The heat of polymerization can be measured by DSC method, measuring thermal effect of polymerization and ignoring the heat produced from decomposition of the initiator and heat of termination. In isothermal experiments sample is placed at a chosen temperature and thermogram is recorded versus time. Assuming typical relationship... 

It has been foimd that for many monomer-template-solvent systems, the rate of reaction is proportional to the initiator concentration in power n and monomer concentration in power m, as in simple radical polymerization (qv). However, exponents n and m in the presence of template are different in comparison with the same system without the template. 

A more recent development is template polymerization [520 522]. When acrylic acid was polymerized in aqueous solution using potassium persulfate as initiator, the polymerization proceeded very slowly. In the presence of poly(vinylpyrrolidone) but under otherwise identical reaction conditions, the rate of polymerization increased dramatically, depending on the amount of PVP. At nearly equimolar concentrations of PVP and monomer, the rate of polymerization reaches a maximum value, because of the strong interaction between poly(vinylpyrrolidone) and acrylic acid in aqueous solution (Scheme 40) [523]. 

The initial rate of polymerization by AMV RTase in vitro is 300 nt/min, a rate nearly equal to that of E. coli DNA Pol I on some templates (9,50). The rates of DNA synthesis by RSV RTase in vivo and in the endogenous reactions have been reported to be 30 nt/min (59), while the rate of DNA synthesis on the poly(A) template by HIV-1 RTase is 60-90 nt/min (60). 

Cohen presented a method for determining the rate constants for irreversible polymerization where the different rate constants apply to the initial step and the propagation reactions. Data are obtained in the usual manner at moderate concentrations of primer. The approach is equally valid for template-directed and template-independent polymerization. Cohen applied this method to obtain rate constants for polyadenylate [(poly(A)] polymerase (EC 2.7.7.19). The report also contains useful information about simulating the time course of irreversible polymerization. 

Using copolymer maleic anhydride/styrene instead of poly(maleic anhydride) homopolymer as a template, the authors found that polymerization of 4-vinyl pyridine proceeds also without any initiator, but with lower rate of reaction. 

Apparent K and kcat values are determined by modifying the nucleotide incorporation assay described above such that variable concentrations of dNTPs and the primed M13 template are employed. To determine the DNA parameters, concentrations of dCTP, dGTP, and dATP are fixed at 200 p,Af and TTP is typically present at 100 p,Af (10 pAf [ HJTTP, 90 pJlf TTP). Polymerization rates are measured in triplicate in the presence of seven or eight different concentrations of the primed M13 template, chosen to bracket the expected Km (see Tables II and III). For the dNTP parameters, the final M13 concentration is typically fixed at 15 nM and polymerization rates are measured at dNTP concentrations ranging from 1 to 200 p,M each dNTP (equimolar concentration of all four dNTPs). Reactions are initiated with the addition of enzyme and allowed to proceed at optimal temperature for up to 15 min. Aliquots are withdrawn at various times and spotted... 

Using polysarcosine with a different DP as an initiator, and a template, of polymerization of DL-phenylalanine NCA, the authors found that the rate of reaction was much faster than those initiated by conesponding low-molecular-weigbt amines. Moreover, it was found tbat tbe rate of polymerization depended on the DP of polysarcosine and on the solvent used. The replacement of phenylanyl emits for sarcosyle units in the polypeptide initiator lowered the initiator efficiency. 

An oxidative polymerization of aniline in the presence of poly(2-acrylamido-2-methyl-l-propanesulfonic acid) (PAMPSA) as a template was described in Reference 124. The process was carried out in water, and ammonium persulfate was used as initiator. The synthesis was performed at room temperature. In similar conditions, but with benzenesulfonic add instead of AMPSA, polymerization of aniline does not occur. According to the authors, the localization and protoni-zation of aniline takes place along the chain of AMPSA. Adding sodium chloride to the reaction solution, considerably decreased in both the process rate and the polyaniline yields. What was interesting was that the system remained phase-homogeneous on all stages of conversions. Kinetics examination lead to the condusion that the process is of a pronounced autocatalytic character. Electroconductivity of the film formed from polyaniline-PAMPSA complex has rather low conductivity of about 10 S cm . ... 

Certain secondary structures in an RNA template may force the RTase to halt and/or terminate reverse transcription, which is often the cause of inefficient cDNA synthesis. To disrupt potential secondary structures, RNA templates are customarily denatured either by brief heating (30-60 sec at 100°C) or by treatment with methylmercuric hydroxide (CHjHgOH, 2.5 mM) followed by neutralization of the reagent with at least three times excess 2-MSH immediately prior to the addition of RTases (30). Despite the initial disruption of secondary structures hy either of the two methods mentioned earlier, RNA templates tend to regain, at least partially, their original secondary struaures during the period of incubation. The elevation of the reaction temperature, sometimes up to 55°C with AMV RTase, helps retard the process of refolding, while the elevated temperature increases the rate of polymerization by the RTase. However, the optimal temperature has to be compromised with the stability of the individual RTase. 

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